vitamin-k-semiquinone-radical and cumene-hydroperoxide

Ca2+ retention in mitochondria, opening of the Cysclosporin A- sensitive permeability transition pore and cell death were studied in Ehrlich ascites tumour cells in the presence of different prooxidants. Low concentrations (1-20 microM) of the prooxidants (menadione, cumenehydroperoxide, t-butylhydroperoxide) induced pore-opening in permeabilized cells at threshold Ca2+ load. Incubation of cells with low concentrations of prooxidants was able to induce cell cycle disturbance and cell death. Under the prooxidant effect, mitochondrial membrane potential drop and Ca2+ retention decrease in mitochondria were found to precede death of Ehrlich ascites tumour cells.

Topics: Animals; Benzene Derivatives; Calcium; Carcinoma, Ehrlich Tumor; Cell Death; Cell Survival; Intracellular Membranes; Membrane Potentials; Mice; Mitochondria; Oxidants; Oxidative Stress; Permeability; Peroxides; tert-Butylhydroperoxide; Tumor Cells, Cultured; Vitamin K

Selenium-dependent glutathione peroxidase (GPx) plays a protective role in oxidative stress-induced apoptosis. In this study, we demonstrated that MDBK cells, a bovine renal epithelial cell line, exhibited internucleosomal DNA fragmentation characteristic of apoptotic cell death under selenium-deficient conditions with lower doses of hydrogen peroxide (H2O2) than under selenium-supplemented ones. This was due to a decreased amount of GPx in the cells under selenium-deficient conditions, because other antioxidative enzyme activities were not affected by the selenium supplementation. Cumene hydroperoxide also induced DNA fragmentation in selenium-deficient cells but no ladder formation was observed. Flow cytometric analysis showed that selenium-deficient cells were less capable of scavenging intracellular peroxides after exposure to exogenous H2O2 than selenium-supplemented ones. In contrast, there was no difference in viability between selenium-supplemented and non-supplemented cells in cell survival after exposure to menadione, which activates the electron transport system and increases intracellular superoxide radicals. Clofibrate, a peroxisomal proliferator and an inducer of catalase (CAT), partially protected both Se-deficient and Se-supplemented cells from exogenous H202. We concluded that selenium-deficient cells were more easily brought to apoptotic cell death by peroxides, but not by superoxide radicals, than selenium-supplemented ones and that CAT could compensate for the depletion of GPx to a certain degree by scavenging H2O2.

Topics: Animals; Antioxidants; Apoptosis; Benzene Derivatives; Catalase; Cattle; Cell Line; Cell Survival; Clofibrate; Glucose Oxidase; Glutathione Peroxidase; Hydrogen Peroxide; Oxidative Stress; Peroxides; Reactive Oxygen Species; Selenious Acid; Selenium; Selenium Compounds; Superoxide Dismutase; Superoxides; Vitamin K; Xanthine Oxidase

The role of reactive oxygen species in injury to cells lining the gastrointestinal tract has gained importance in recent times, and their role in intestinal diseases has been studied.. In this study colonic epithelial cells isolated from the rat colon were exposed in vitro to various oxidants such as menadione, xanthine-xanthine oxidase, hydrogen peroxide, cumene hydroperoxide, and tertiary butyl hydroperoxide, separately. Changes in cell viability, thiol status, and the antioxidant enzyme activities were measured.. Colonocytes were found to be sensitive to menadione and were not affected by various other oxidants. Decrease in cell viability, depletion of reduced glutathione and protein thiol, and change in antioxidant enzyme activities were observed when the cells were exposed to menadione.. This study suggests that, unlike other cells, colonocytes are susceptible only to certain selective oxidants.

Topics: Animals; Benzene Derivatives; Catalase; Cell Survival; Cells, Cultured; Colon; Epithelial Cells; Epithelium; Glutathione; Glutathione Peroxidase; Glutathione Reductase; Glutathione Transferase; Hydrogen Peroxide; Oxidants; Peroxides; Rats; Sulfhydryl Compounds; tert-Butylhydroperoxide; Vitamin K; Xanthine; Xanthine Oxidase; Xanthines

The role of seleno-glutathione peroxidase (GSHPx; EC 1.11.1.9) in the cellular defense against oxidative stress was selectively investigated in novel cell models. Expression vectors designed to overexpress human GSHPx efficiently in a broad range of mammalian cells were used to transfect T47D human breast cells which contain very low levels of endogenous GSHPx. Several stable transfectants expressing GSHPx to various extents, up to 10-100 times more than parental cells, were isolated and characterized. Growth inhibition kinetics following transient exposure to increasing concentrations of H2O2, cumene hydroperoxide or menadione (an intracellular source of free radicals and reactive oxygen intermediates) showed that transfectants overexpressing GSHPx were considerably more resistant than control T47D cell derivatives to each of these oxidants. A sensitive DNA end-labeling procedure was used as a novel approach to compare relative extents of DNA strand breakage in these cells. In contrast to the extensive DNA damage induced in control transfectants by 1-h exposure to cytotoxic concentrations of menadione, the extent of DNA breakage detected in GSHPx-rich transfectants was remarkably reduced (6- to 9-fold, p less than 0.005).

Topics: Benzene Derivatives; Blotting, Northern; Blotting, Southern; Blotting, Western; Breast Neoplasms; Cell Division; DNA; DNA Damage; Gene Expression; Glutathione Peroxidase; Humans; Hydrogen Peroxide; In Vitro Techniques; Mutagens; Oxidants; RNA, Messenger; Transfection; Vitamin K

Biliary epithelial cells (BEC) and parenchymal cells isolated from normal rat liver were exposed in vitro to a number of toxic compounds. BEC were found to be highly sensitive to concentrations of menadione (100 microM) and cumene hydroperoxide (10 microM) that are usually not effective as toxic agents in hepatocytes under normoxic conditions. On the other hand, BEC were not affected by concentrations of carbon tetrachloride or 7-ethoxycoumarin that are known to exert toxic effects on hepatocytes. The development of both menadione- and cumene hydroperoxide-induced toxic injury in BEC followed a common and time-correlated pattern, and included a strong depletion of GSH, depletion of protein thiols and an increase in the extent of cell death. The damage induced by cumene hydroperoxide was found to be independent of lipid peroxidative processes and was prevented by a pre-incubation with desferrioxamine. The cytotoxicity of menadione was further exacerbated by dicoumarol but was not prevented by desferrioxamine or promethazine. The mechanisms underlying BEC injury and death induced by the quinone and by the organic hydroperoxide are discussed in relation to the known biochemical characteristics of BEC.

Topics: Animals; Benzene Derivatives; Carbon Tetrachloride; Cell Survival; Cells, Cultured; Deferoxamine; Epithelial Cells; Epithelium; Glutathione; Glutathione Reductase; Liver; Male; Oxidation-Reduction; Rats; Time Factors; Vitamin K

Menadione (2-methyl-1,4-naphthoquinone) is a remarkably potent inhibitor of microsomal lipid peroxidation, effective at submicromolar concentrations. Its possible mechanism of action and the relationship between naphthoquinone structure and antioxidant activity were the topics of this investigation. In the microsomal lipid-peroxidizing system dependent on NADPH and ferric pyrophosphate, menadione, at concentrations of 50 microM or higher virtually eliminated the accumulation of malondialdehyde and lipid hydroperoxides. In the NADPH-independent, cumene hydroperoxide-dependent system, menadione was also an effective antioxidant, but only in the presence of reducing equivalents. These and other observations indicate that a reduced form of menadione, either the hydroquinone or semiquinone, is the active antioxidant, and suggest that it may trap hydroperoxy radicals, alkoxy radicals, or other free radicals involved in propagating lipid peroxidation. Moreover, these results show that electron diversion per se cannot account for the antioxidant effects of menadione. A comparison of the antioxidant activities of eight 1,4-naphthoquinones indicated that methyl substitution of C-2, lack of steric hindrance at C-3 or C-5, and (in the case of weak acids) a relatively high pKa are favorable structural features associated with strong antioxidant activity.

Topics: Animals; Benzene Derivatives; Hydrogen-Ion Concentration; Iron; Lipid Peroxides; Male; Microsomes, Liver; NADP; Naphthoquinones; Rats; Structure-Activity Relationship; Vitamin K

Isolated rat liver cells have been exposed to 3 different lipid peroxidation-inducing agents, CCl4, FeCl3 and cumene hydroperoxide, and the rates of malonaldehyde production and of lipoprotein secretion have been compared. Results indicate that it is possible to induce a high degree of lipid peroxidation without inducing strong changes in lipoprotein secretion. Only in CCl4-poisoned hepatocytes is lipoprotein secretion strongly impaired. In this experimental condition, the effect of free radical scavengers, or inhibitors of lipid peroxidation, as well as the behavior of both lipid peroxidation and lipoprotein secretion, have been evaluated. Promethazine and propyl gallate prevented malonaldehyde production, but neither agent reduced covalent binding nor improved secretion. Menadione, on the contrary, besides inhibiting malonaldehyde production, decreased covalent binding and protected against the impairment of secretion. These data lead to the conclusion that covalent binding of CCl4 metabolites, rather than lipid peroxidation products, accounts for the derangement of lipoprotein secretion in CCl4-poisoned liver cells.

Topics: Animals; Benzene Derivatives; Carbon Tetrachloride Poisoning; Chlorides; Ferric Compounds; Lipid Peroxides; Lipoproteins; Liver; Male; Malondialdehyde; Palmitic Acid; Palmitic Acids; Promethazine; Rats; Rats, Inbred Strains; Vitamin K